Pndmoni ile enfekte hastalarda Klebsiella pntimoni'nin immunomodulatorii olarak miRNA‐ 155 ve miRNAa‐23'in rolü

Klebsiella pneumoniae, is an opportunistic bacteria that infected immunocompromised patients, and to evaluate the effectiveness of micro ribonucleic acid (miRNA-23 and miRNA-155) as biomarkers for detecting K. pneumoniae infection in patients serum. this study aims to isolate and diagnose K. pneumoniae from patient as well as to provide the latest understanding of interleukins and molecular pathogenesis. One hundred and fifty samples (sputum, lung fluids, and blood) were collected in sterile containers from patients attending two hospitals at Baghdad Teaching Hospital, National Tuberculosis Institute, Baghdad - Iraq for the period from October 2022 to March 2023. Seventy-five of K. pneumoniae isolates were verified from 150 specimens using 75 samples of patients' serum to examine the immune and molecular levels. After confirming the diagnosis of bacteria by microscopic, biochemical assays, and the VITEK® 2compact system. The bacteria were found to be fermenting lactose on the macckonkey agar and were negative for the oxidase test but positive for the catalase test. Patients were divided into four age groups; one group (age 25–35), group two (age 36–45), group three (age 46–55), and group four (age 56–70) with apparently healthy controls matching with age and gender were collected 75 blood samples from people who donate to the Iraqi National Blood Bank. The level of interleukins in the serum of the patient and the healthy group was also evaluated using ELISA. The healthy control group had a lower mean value of serum IL-1ꞵ levels (86.75 ± 2.31) pg/mL compared to (345.14 ± 7.18) pg/mL and IL-34 level in pneumonia patients showed a significant increase (154.48 ± 15.8) pg/ml compared to the control (74.51 ± 1.55) pg/ml. There was also a significant increase (P< 0.01) in serum TLR4 level in patients (10.17 ± 0.22) ng/ml compared to control (2.38 ± 0.06) ng/ml. According to the receiver operating characteristic (ROC) curve results, true positive results for IL-1ꞵ, IL-34, and TLR4 were observed. ROC and area under the curve (AUC) were used to evaluate the diagnostic power of IL-1ꞵ, IL-34, and TLR4 receptors as potential biomarkers for pneumonia patients compared to healthy controls. MiRNA-23 and miRNA-155 were also recognized by using RT-PCR and found the pathogenicity of K. pneumoniae differed, according to the individual results of the relative expression level ∆CT of miRNA-23 and miRNA-155 in each of the four groups that were found in the serum of patients who had previously been infected. Regarding miRNA-155 ∆CT (8.334 ± 2.193 and -3.518 ± 0.618, respectively) and the mean of the healthy controls (1.169 ± 1.669 and -7.475 ± 0.698, respectively), only groups one and two showed a significant difference in relative expression levels in the patient's serum Mean ± S.E. A significant difference was observed in the ∆CT value only in group two, involving patients' expression of miRNA-23 was roughly (9.544 ± 0.749) in comparison to the control group's (4.689 ± 0.763). The analysis determined the fold change 2-(∆∆C T ) of the miRNA-23 and miRNA-155 genes, which exhibited changes in response to their triggered levels of expression against K. pneumoniae. Furthermore, K. pneumoniae was more effective in significantly increasing the expression levels of miRNA-23 and miRNA-155 in patients' serum in group one, as indicated by the fold changes 2-(∆∆C T ) test of the miRNA-23 and miRNA-155 genes in the current study. This test showed a significant difference between the miRNA-23 and miRNA-155 genes in group one by score (10.52 and 5.65%, respectively). Also, the study reveals low levels of miRNA-23 and miRNA-155 expression in groups two, three, and four 2.501, additionally, the fold change 2 -(∆∆C T ) analysis did not demonstrate any significant differences.The comparison assay between Mean ± S.E. of fold changes 2 -(∆∆C T ) of miRNA-23, miRNA-155, and Mean ±S.E. of interleukins IL-1β, IL-34, and TLR4 was done to find positive correlations and significant P-values. The results of group one observed a negative correlation between miRNA-23 and IL-1β, IL-34, TLR4 in contrast to miRNA-155 which had a positive correlation only with TLR4 (0.511), and the difference was extremely significant (P<0.01) between IL-1β, IL-34, TLR4 and both miRNA-23 and miRNA-155. Furthermore, group two data showed a high positive correlation between miRNA-23 and IL-1β, IL-34 and TLR4 (0.983, 0.969, and 0.376, respectively),whereas miRNA-155 exhibited positive correlations with IL-1β and IL-34 (0.982 and 0.966, respectively). There were significant differences between IL-1β and IL-34, as well as the genes miRNA-23 and miRNA-155 (P< 0.01). According to the data of the third group, miRNA-23 and miRNA-155 correlated positively with IL-1β and IL- 34 by (0.959, -0.527) and (0.958, -0.440), respectively, and both genes appeared a highly critical difference (P< 0.01) with only these two interleukins.The data of group four demonstrated positive correlations between miRNA- 23, miRNA-155, and IL-34, TLR4 with values of (0.983, 0.668), and (0.982, 0.427), respectively. Significant differences were found between IL-1β, IL-34, TLR4, and miRNA-23, miRNA-155 at both P-values (P< 0.01) and (P< 0.05). The study is evaluating the efficacy of interleukins IL-1β, IL-34, and TLR4, and genes miRNA-23 and miRNA-155 as biomarkers for the early detection of K. pneumoniae infections in patients. Patients in groups two, three, and four with low levels of 2-(∆∆C T ) miRNA-23 and miRNA-155 in gene expression may be at higher risk of K. pneumoniae infection. the exposure may affect circulating miRNAs implicated in biological processes related to pneumonia, including immune function. All patients in group one exhibited significant differences, with higher fold changes 2 -(∆∆C T ) in the levels of miRNA-23 and miRNA-155 gene expression. These biomarkers hold potential for the early identification of K. pneumoniae infections.